Project 2 Project Leader: Yazaki, Paul Principal Investigator: RaubitSChek,Andrew DESCRIPTION: To enhance the therapeutic efficacy of radioimmunotherapy, pretargeting strategies have been developed to uncouple the antibody's slow tumor targeting and clearance properties from the delivery of a radiotherapeutic agent. Separation of these two steps allows for the rapid delivery of a therapeutic dose reducing the exposure to normal tissues, especially the bone marrow. Although previous pretargeting strategies have demonstrated clinical responses, major challenges have blocked their clinical utility. Problematic issues include immunogenicity of tumor targeting agents, healthy normal tissue exposure to the radiotherapeutic agent, and purity and production of reagents. Nonetheless, these challenges offer tremendous opportunities for the improvement of an efficient multi-step delivery system. The carcinoembryonic antigen (CEA) system remains the primary focus of our work. In this project, the proposed work includes three approaches to address these issues. The first specific aim is a logical extension of the clinically tested three-step biotin-based approach, incorporating an anti-biotin antibody as the bridging agent between biotinylated anti-CEA antibody and biotin-^Y-DOTA. The second is the development of a bispecific antibody approach that targets both CEA-positive tumors and the 9CY-DOTA radioligand. The well characterized anti-CEA T84.66 and anti-DOTA 2D12.5 monoclonal antibodies will be designed into recombinant bispecific antibodies based on a bispecific diabody, tandem diabody and di-diabody-Fc format. The third aim explores a completely new approach made possible by the discovery of an exciting single variable domain antibody-based scaffold technology that will allow the production of novel anti-CEA and anti- CEA/CEA complex targeting agents. These aims will be conducted in parallel but are not exclusive in their pathways. Knowledge gained from all aspects of the Project will be incorporated toward the development of a single pretargeting system for clinical application.